Summary Breast cancer has become a curable disease if it is diagnosed at an early stage; however, more than 90% of breast cancer deaths are still attributed to metastases and bone is the major distant organ of metastatic breast tumor growth. Therefore, it is of paramount importance to elucidate the exact pathological mechanism of bone metastasis to identify a specific therapeutic target for this devastating disease. The recent cancer stem cell theory, which still remains as a hypothesis, predicts that there is a distinct population of metastatic cells that have stem-like characteristics as well as invasive ability. Our preliminary data indicate that (i) cancer stem-like cells (CSCs) from highly metastatic cell lines express a high level of hyaluronan synthase gene 2 (HAS2) which is significantly correlated with patient metastasis-free survival, (ii) HAS2 in CSCs significantly activates the expression of PDGF-BB in tumor associated macrophage (TAM) through interaction of hyaluronic acid (HA) in CSCs and CD44 in TAM, which then activates the expression of FGF7 and FGF9 in bone niche cells followed by stimulating self-renewal of CSCs and (iii) the inhibitor of HAS2, 4MU, can block this vicious cycle in the bone niche and significantly suppress bone metastasis in vitro and in vivo. In the current grant application, we propose our novel hypotheses that (i) high expression of HA in CSCs facilitates a take-over of pre- existing niche of hematopoietic stem cells (HSC) in the bone and activates TAM and mobilizes the bone niche, which in turn promotes self-renewal of CSC and (ii) a prodrug inhibitor for HAS2 can be used for preventive and therapeutic intervention of metastatic disease. To test these hypotheses, we will first examine the role of HAS2 of CSCs in bone metastasis in vivo and also test the efficacy of a novel pro- drug of HAS2 inhibitor in our animal model of bone metastasis (Aim 1). We will also elucidate the mechanism by which HAS2 promotes self-renewal of CSC by mobilizing bone microenvironment in vitro (Aim 2). Furthermore, we will directly examine the clinical relevance of HAS2 in breast cancer metastasis and identify potential diagnostic/prognostic markers for bone metastasis (Aim 3). The ultimate goal of this project is to define the pathological process of bone metastasis and identify specific therapeutic and preventive targets for this devastating disease. We do believe that the proposed hypothesis, if proven to be valid, will present a novel paradigm to understand the pathological mechanism of bone metastasis which will significantly impact the treatment of breast cancer.